Hepatitis C virus (HCV), a pathogen that infects over 150 million people worldwide, is the leading cause of chronic hepatitis, liver cirrhosis and hepatocellular carcinoma. HCV is a genetically diverse virus with 6 known genotypes with genotypes 1 and 3 being the most prevalent. This genetic diversity makes HCV infection difficult to treat. In the last few years, the advent of direct-acting antivirals (DAAs) has remarkably improved therapeutic options and treatment outcomes. However, despite highly potent inhibitors against multiple proteins, drug resistance is a major problem in all drug classes. Drug resistance is a loss of inhibitor potency while maintaining substrate processing. Though NS3/4A protease inhibitors are highly potent, they are not efficacious against all genotypes and are susceptible to drug resistance. Underlying differential inhibitor potency are the molecular mechanisms of drug resistance and genotypic differences. Elucidating these are key to developing protease inhibitors that avoid drug resistance and are effective against all HCV genotypes. Specifically most protease inhibitors in clinical development contain P2 moieties that contact unessential residues of the protease, which while increasing potency also increases their susceptibility to single site mutations that confer drug resistance. I hypothesize that protease inhibitors that avoid contact with these residues while leveraging contact with unexploited areas in the active site will result in inhibitors with enhanced potency and higher barriers to drug resistance. To investigate this hypothesis, using computational techniques, I will design a panel of novel protease inhibitors with extended P4 groups. I will then synthesize and enzymatically assay these protease inhibitors. Top leads will be co-crystalized with the protease and structurally analyzed to optimize the computational designs and initiate iterative rounds of inhibitor design. This project will provide molecular insights about the mechanisms of drug resistance as well as new strategies for the design of novel protease inhibitors for the effective treatment of HCV infection.